Abstract: A method of liquid chromatography tandem mass spectrometry (LC-MS/MS) was developed for the determination of arachidonic acid (AA), 13-hydroxyoctadecadienoic acid (13-HODE) and 9-hydroxyoctadecadienoic acid (9-HODE) in plasma．The plasma sample was extracted using solid phase extraction, then separated on BEH C18 column with equal elution of acetonitrile and water as mobile phase, and the flow rate was 0.2 mL/min. It was detected by electrospray ionization tandem mass spectrometry in the multiple reaction monitoring negative mode. The results show that AA, 13-HODE and 9-HODE have good linear relationship in the concentration of 0.5-50 μg/L. The average recoveries are 97.42%-101.46% at 3 spiked levels. The precisions of intra-day and inter-day are less than 6.4%. The limits of quantification are 0.5, 0.5, 1.0 μg/L for AA, 13-HODE and 9-HODE, respectively. The method is simple, efficient and accurate, which is suitable for determination of fatty acids and metabolites in plasma.